The long term goal of this project is to determine the mechanisms by which ischemia and reperfusion injure the isolated lung and whether these mechanisms are relevant to ischemia-reperfusion lung injury in intact animals. We hypothesize that unlike other organs the lung is injureed by toxic oxygen metabolites generated during ischemia, when oxygen tension is high and glucose depleted. The endothelium is a primary target of injury, which, therefore, results in increased vascular permeability and edema formation upon reperfusion. To test this hypothesis, experiments will be performed in isolated sheep lungs to quantitate the effects of ischemic duration and oxygen tension, confirm that the O2- dependence of the injury is related to production of toxic O2 metabolites, and determine the source of these metabolites. Parallel experiments using cultured sheep pulmonary endothelial cells in an in vitro model of lung ischemia-reperfusion will evaluate endothelium both as a target and as a source of injury. Isolated lungs and pulmonary endothelial cells from dogs will be studied to confirm that ischemia-reperfusion lung injury is less severe in this species and to determine the mechanisms of the difference. The response and role of the bronchial circulation will be evaluated in vivo and in isolated lungs from sheep, a species anatomically suited to this purpose. Hemodynamics, fluid balance, antioxidant activity and energy state will be measured in isolated lungs. Viability, biochemical function, and barrier function will be measured in endothelial cells. In addition, free radical concentrations will be measured in both preparations by eletron paramagnetic resonance spectroscopy to assess directly the role and identify of these substances. Electron microscopic evaluation of endothelium in lung and monolayer will be performed to establish structure-function correlations. Therapeutic interventions suggested by the above studies will be tested in intact animals acutely after transient left lung ischemia and chroncially after left lung autotransplantation. Preliminary experiments confirm the feasibility and usefulness of these approaches. The results could have significant implications with respect to human conditions in which lung ischemia-reperfusion is thought to play a pathogenetic role, such as cardiopulmonary bypass, pulmonary thromboendarectomy and thrombolysis after pulmonary embolism, and lung transplantation.
